ISO 18337:2015

Overview

ISO 18337:2015 specifies a practical method to measure the lateral resolution of a confocal fluorescence microscope (CFM). The standard describes imaging an object much smaller than the expected resolution and analysing the resulting point spread function (PSF) to determine the lateral resolution, typically expressed as the full width at half-maximum (FWHM) of the PSF. The procedure is written to be suitable for non‑expert operators and supports consistent performance assessment for CFMs used in surface chemical analysis and imaging of thin films and nanomaterials.

Key topics and technical requirements

• Measurement principle: Image a sub‑resolution fluorescent object so the recorded image represents the convolution of the PSF and the object; estimate lateral resolution from the PSF FWHM.
• Sample selection: Use very small fluorescent particles (e.g., polymer beads, quantum dots). ISO 18337:2015 notes that a particle smaller than one‑fourth of the PSF contributes <1% to the observed size, avoiding the need for deconvolution.
• Data extraction: Select an appropriate spot and extract a line profile (band‑average process) to compute the PSF FWHM.
• Instrumental parameters affecting lateral resolution:
  • Objective lens (OL) and numerical aperture (NA) - primary determinants of focus size.
  • Detection pinhole size and tube lens focal length - influence confocality and apparent pinhole image on the sample plane.
  • Laser beam collimation and purity, polarization, and excitation/emission wavelengths - these alter PSF shape and size.
  • Image contrast - detector sensitivity, optics cleanness, and signal‑to‑noise impact measurement accuracy.
• CFM operation types covered: stage‑scanning, laser‑scanning, and spinning‑disk configurations, with guidance on practical differences affecting resolution.

Applications

• Performance verification and acceptance testing of confocal fluorescence microscopes.
• Routine quality control in microscopy core facilities and instrument manufacturers.
• Comparative benchmarking of CFMs for imaging thin films, nanomaterials, and fluorescently labelled samples.
• Supporting surface chemical analysis workflows where spatial resolution impacts interpretation of compositional maps.

Who should use this standard

• Microscopy facility managers and technicians performing instrument qualification.
• Engineers and scientists validating CFM performance in research and industrial labs.
• Instrument manufacturers and service engineers for calibration and specification verification.
• Quality assurance and accreditation bodies assessing laboratory imaging capability.

Related standards

• ISO 18337:2015 references ISO 18516 for related terms and analysis procedures. Users should consult these documents together for comprehensive surface chemical analysis and PSF characterisation guidance.

Keywords: ISO 18337:2015, confocal fluorescence microscope, lateral resolution, PSF, FWHM, fluorescent nanoparticles, quantum dots, objective lens, numerical aperture, detection pinhole, imaging.